A library of bacteria: students categorizing E. coli outbreaks

Cal Poly students and faculty are working together to build a library of DNA fingerprints to identify the culprit when people get sick from E. coli. In 2007, health officials posted notices in Pismo Beach because of a high increase in E. coli levels, biological sciences chair Christopher Kitts said.

E. coli is a bacterium that comes from a variety of sources, but it’s found in the same place. The bacterium is present in the guts of humans and animals, said Anya Goodman, chemistry and biochemistry professor.

“E. coli is just kind of used an indicator of fecal contamination, because normally people don’t just expect it to be found in the environment as a free living thing,” Goodman said. “It’s bacteria that lives in the gut.”

Identifying the source of E. coli can get complicated. Kitts, faculty and students are researching how to better identify the source of E. coli to reduce its presence in local waters.

Sometimes the bacteria pass from one animal to another and then into the water, Goodman said.

“Part of the problem is they can be interchangeable,” Goodman said. “So I’ve heard stories that there’s exchange of E. coli between seagulls feeding on trashed, poopy diapers. So part of what we’re trying to figure out is what are some of the host specific E. coli and can we use them to track the sources of contamination.”

Just as each person has a different fingerprint, each E. coli is a little different from species to species, Goodman said.

“It’s really easy to say ‘this is E. coli or ‘not E. coli,’ but it’s really hard to distinguish different E. coli’s from each other,” Goodman said.

The team is researching to find a simpler, yet accurate method of testing and identifying E. coli sources, Kitts said. E. coli is E. coli, but bird E. coli will have a different molecular fingerprint than dog E. coli.

Students in biology professor Michael Blacks’ BIO 161, Introduction to Cell and Molecular Biology, class are analyzing samples of E. coli to add fingerprints to the library. Without the initial sample, the library wouldn’t have data to compare E. coli to.

“The fingerprinting method that we have developed here is novel,” Black said.

The method was developed by a team of faculty members at Cal Poly and is being practiced by students in freshmen and sophomore biology classes.

“Pyroprinting is a method that allows us to determine the sequence of a DNA strand,” Black said. “DNA is based on four different letters which represent nucleotides that make up the strand. As enzymes incorporate a letter, a pyrophosphate is released as a byproduct of the reaction. Other enzymes in the mix use this pyrophosphate to convert chemical energy into light energy — the same enzymes that make fireflies glow. This means that each times a letter is added, light is given off and can be detected by the machine.”

The National Science Foundation initially turned Cal Poly down for a grant because there was a lot of mistrust in the accuracy of the students’ work. Cal Poly demonstrated that not only can undergraduate students collect, properly isolate and test a sample for E. coli, but that they can perform the same tests and produce the same results with very little variation, Goodman said.

“We designed a study where there are triplicate repeats, so three students do the same thing and we can show ‘Yeah, they can do real science. They can do it.’ That’s partly why we ended up getting this grant,” Goodman said. “The first time there was a lot of mistrust and we said, ‘Look, we’re doing it. Our students are that good, they can handle this.’”

With the new method for analyzing DNA, it’s possible for even freshman-level undergraduate students to perform the same type of tests that are conducted in large testing laboratories.

“We can distinguish between very closely related bacteria,” Black said.

While the fingerprint library is still a work in progress, it gives students an opportunity to practice real world techniques and be a part of a newly evolving method for identifying and sequencing bacteria DNA, Goodman said.

“The tools they’re using and the statistical analysis that they’re using and the exposure to database type research is all very valuable and applicable to a variety of different things,” Black said.

Professor Janelle Barbier’s students in General Microbiology I (MCRO 224) are testing samples collected by other biology classes and isolating E. coli to be analyzed and added to the library.

“Learning microbiology techniques not only helps build the library, but these are all standard tests and are used in a variety of disciplines” Barbier said.

Doing real world work gives Cal Poly students an advantage, Black said. Putting the research into the fingerprint library will help scientists pinpoint the source of contamination.

“If we can figure out the source of the E. coli, we can halt that transmission, if possible,” Black said.

Megan Stone contributed to this article.